Peptide synthesizers and polynucleotide synthesizers have a source of building blocks, which are, respectively, amino acids and nucleotide bases, that are sequentially supplied to a reactor column where they are chemically reacted to form a peptide or an oligonucleotide. Such a reaction requires carefully controlled conditions in which the presence of an incorrect amino acid (or nucleotide base, respectively), leads to the synthesis of the wrong peptide or an incorrect oligonucleotide. As a result, each stepwise addition is followed by a washing of appropriate flow lines. Although the washing delays the process, the delay is minimized by minimizing the volume of the flow lines involved.
A further aspect of the problem is the method in which the amino acids for the peptide, or nucleotide bases for the oligonucleotide, are supplied from their containers. Positive gas displacement is preferred, inasmuch as any system that attempts to "pull" them out using a vacuum, runs the risk of generating bubbles in the liquid, since the liquid amino acids or nucleotides are not degassed prior to use. However, such positive gas displacement systems in turn render downstream control of the displaced liquid difficult.
DNA synthesizers have been constructed to overcome such a control problem and retain the use of gas displacement of the bases. To allow the bases to flow into the system, including a reactor column, at a controlled rate and volume, a syringe controlling means, also called a controller, has been positioned between the reactant (hereinafter, a "base") outlet line, fed from an 8-port rotary valve, and the reactor column. When the syringe piston is withdrawn by a motor to create a storage chamber, the positive gas pressure on the supply bottles causes one or more bases to feed into the syringe. Thereafter, the syringe piston is reversed and the bases are injected into the reactor column. An example of such a conventional synthesizer is the Autoinject module of the Cruachem DNA synthesizer.
A problem with such an instrument is that the syringe becomes "contaminated" with the bases that are to be delivered to the reactor column, due to its upstream location. Such a condition requires that the syringe be thoroughly cleaned (with acetonitrile) prior to the drawing of the next base in the sequence, since the presence of the wrong base will ruin the DNA sequence. Because of the relatively large volume of the syringe, such cleaning is time-consuming and difficult. Yet, if cleaning is minimized, contamination will occur.
Yet another problem with such instruments is that the syringe controller is not in a position to control the feed of the reagents, such as iodine and the like, needed in the synthesis, since the reagents are fed into the reactor via a line separate from the line supplying the bases.
Still another problem with such a synthesizer has been the use of an 8-port rotary valve to collect the bases sequentially. As the valve wears, it leaks, and the base sequence is no longer free of contamination from other bases. To deal with this problem, parallel manifolds have been suggested as in U.S. Pat. No. 4,598,049. However, the manifolds taught therein do not provide for separate washing of each base inlet in sequence, so that there is the chance that one or more base inlets will be inadequately washed, and contamination will occur.
Thus, prior to this invention there has been a need for a DNA synthesizer that is free of these problems.